Adverse birth outcomes and associated factors among newborns delivered in Sao Tome & Principe: A case‒control study

Background Newborns with one-or-more adverse birth outcomes (ABOs) are at greater risk of mortality or long-term morbidity with health impacts into adulthood. Hence, identifying ABO-associated factors is crucial for devising relevant interventions. For this study, ABOs were defined as prematurity (PTB) for gestational age <37 weeks, low birth weight (LBW) <2.5 kg, macrosomia >4 kg, asphyxia for a 5-minute Apgar score <7, congenital anomalies, and neonatal sepsis. This study aimed to assess factors associated with ABOs among babies delivered at the only hospital of Sao Tome & Principe (STP), a resource-constrained sub-Saharan-Central African country. Methods A hospital-based unmatched case‒control study was conducted among newborns from randomly selected mothers. Newborns with one-or-more ABO were the cases (ABO group), while healthy newborns were the controls (no-ABO group). Data were collected by a face-to-face interview and abstracted from antenatal care (ANC) pregnancy cards and medical records. Multivariable logistic regression analysis was performed to identify ABO-associated factors considering a level of significance of α = 0.05. Results A total of 519 newborns (176 with ABO and 343 no-ABO) were enrolled. The mean gestational age and birthweight of cases and controls were 36 (SD = 3.7) weeks with 2659 (SD = 881.44) g and 39.6 (SD = 1.0) weeks with 3256 (SD = 345.83) g, respectively. In the multivariable analysis, twin pregnancy [aOR 4.92, 95% CI 2.25–10.74], prolonged rupture of membranes [aOR 3.43, 95% CI 1.69–6.95], and meconium- fluid [aOR 1.59, 95% CI 0.97–2.62] were significantly associated with ABOs. Eight or more ANC contacts were found to be protective [aOR 0.33, 95% CI 0.18–0.60, p<0.001]. Conclusion Modifiable factors were associated with ABOs in this study and should be considered in cost-effective interventions. The provision of high-quality ANC should be a priority. Twin pregnancies and intrapartum factors such as prolonged rupture of membranes and meconium-stained amniotic fluid are red flags for ABOs that should receive prompt intervention and follow-up.

Introduction [4,18]. Thus, knowing STP context-specific reality of ABOs enables to target and implement the most proper evidence-based interventions for this setting. For this study, ABOs were defined as PTB, LBW, macrosomia, major congenital anomaly, birth asphyxia and neonatal sepsis suspicion. STP has significant health resource-constraints, since there are no blood cultures or other microbiologic techniques available, and other procedures such as umbilical arterial blood gas analysis, are not feasible [22,23]. Moreover, there are no neonatal intensive care units and, similar to other units in LMICs, lacks a mechanical ventilator or any continuous positive airway pressure machine and surfactant therapy for neonatal care [24,25]. Therefore, birth asphyxia is only determined by using the APGAR score [26][27][28][29], and early-onset neonatal sepsis diagnosis is done in a suspicion-based algorithm [24,25].
Thus, this study aimed to improve neonatal health outcomes in this very resource-constrained SSA country by identifying the factors associated with ABOs among newborns delivered at the only hospital maternity unit in STP. Thus, knowledge of the burden of ABOs and key associated factors in this specific setting can be used to design targeted interventions and better allocate resources for effective ABO reduction.

Study design and period
A facility-based unmatched case-control study was conducted in STP among 519 newborns whose mothers gave birth at Hospital Dr. Ayres de Menezes (HAM) Maternity Unit. The recruitment of newborns' mothers occurred from July 2016 to November 2018.

Setting
The archipelago of STP, the smallest Central SSA country, is a 219 161-inhabitant country, with a young population and an annual birth cohort of approximately 6.521 babies [19]. The rate of deliveries in health units is approximately 98%, with 82.4% occurring at the HAM maternity unit, the only hospital in the country [20]. The HAM is a tertiary healthcare facility and receives complicated cases referred from facilities with lower levels of care, as it is the only facility with Comprehensive Emergency Obstetric Care (CEmOC) capable of providing blood transfusions and performing cesarean sections.
The maternity unit has a resource-constrained unit for small and sick newborn babies-Newborn Care Unit (NCU)-with six baby cots, in which babies receive oxygen through nasal prongs or face masks and are assisted by general doctors and nurses, since there are no neonatologists and only two pediatricians in STP.

Participants
The eligibility criteria for participants were as follows: 1) all neonates delivered at HAM and 2) newborns who were born outside the hospital but were later admitted at HAM on the day of birth. A total of 535 newborns were initially enrolled.
The exclusion criteria included the following: 1) all neonates delivered at HAM born without any signs of life (stillbirths), 2) newborns whose mothers had cognitive impairment, and 3) adolescent or illiterate mothers who had not obtained permission from their parents or legal guardians to participate in the study. Sixteen met the exclusion criteria (stillbirths), with a total of 519 participants enrolled.

Selection of cases and controls
Cases (ABO group) were newborns with at least one adverse birth outcome: PTB, LBW, macrosomia, birth asphyxia, congenital anomaly and/or probable neonatal sepsis (see Operational definition of variables). Controls (no-ABO group) were healthy newborns without adverse birth outcomes (� 37 gestational weeks at birth, weight � 2.5 kg at birth and not greater than 4 kg, 5-min APGAR score � 7, no congenital anomaly, and no probable sepsis).

Sample size determination and sampling procedures
Sample size followed the WHO-steps approach [30] applying a web-based sample size calculator, Raosoft, which suggested a minimum sample size of S = 355, which placed the right dimension between 355 (95%) and 579 (99%) confidence [31]. A total of 535 participants were enrolled based on the following assumptions: two-sided 95% confidence level, and power of 80% to detect an odds ratio of at least 2 for ABOs. This sample size was also supported by PASS software [32].
A random sampling was applied to recruit the newborn's mother, selecting every second interval folder from the pile of mothers´medical folders, and occurred during daytime hours of working days. The mothers who consented were interviewed after recovery from the delivery, and the mother-newborn dyads were followed-up throughout their stays until hospital discharge.

Data collection tools, procedures, and quality control
Data were collected by a pre-tested, structured interviewer-administered questionnaire developed from the STP Demographic and Health Survey (DHS) and similar studies [18,19,20]. Additionally, a standard abstraction checklist from ANC pregnancy cards and maternal and newborn records was conducted. The questionnaire contains information related to the sociodemographic characteristics of parents, preconception healthcare services and obstetricsrelated characteristics of the mothers.
To ensure data quality, pretesting was performed on 10 cases and 13 controls and modified based on the findings, mainly adjusting terminology for more culturally friendly terms. The questionnaires were checked for completeness and consistency. Data collection was regularly reviewed by the supervisors. The principal investigator (a pediatrician) executed and was responsible for the field activities as follows: i) obtaining consent and enrollment of the mothers, ii) data collection from ANC cards plus maternal and newborn records, iii) newborns' clinical exams (for diagnosis confirmation), iv) face-to-face interviews, and v) entry of all data collection into the app survey tool. Finally, double data entry was performed to minimize errors during data entry.

Operational definition of variables
ABOs: 1) preterm was defined as a birth that occurred before 37 completed weeks (less than 259 days) of gestation [33]; 2) LBW as a weight of < 2.5 kg at birth [34,35]; 3) macrosomia as a birth-weight over 4000 g irrespective of gestational age [36]; 4) birth asphyxia as an APGAR score at 5-minutes inferior to seven [26,27]; 5) congenital anomaly as structural changes in one or more parts of the infant's body that are present at birth [16]; and/or 6) probable neonatal sepsis or early-onset neonatal sepsis was defined through a clinical-based algorithm as having one or more of the following: i) newborns with early suspicious signs and symptoms (hypothermia or fever, lethargy, poor perfusion, hypotonia, bulging fontanel, respiratory distress, apnea, and gasping respiration), ii) with or without an identifiable maternal infectious risk, and/or iii) requiring NCU admission and antibiotic treatment [37,38].
Gestational age: estimated from the date of onset of the last normal menstrual period or through ultrasound dating of pregnancy [12,40].

Data analysis
Data were entered into the QuickTapSurvey app (2010-2021 Formstack), an offline survey app tool, and exported to Excel to be further checked for completeness, coded, entered, and cleaned. Data were analyzed using SPSS version 25. In this study, cases were coded as 1, and controls were coded as 0 for analysis. The proportion of missing data ranged from 0.8 to 10% across variables. Missing values higher than 10% were described in the analysis.
Descriptive statistics were used to describe the frequency distribution of each of the variables mentioned earlier. The chi-square test was used to compare the proportion of cases and controls between selected categorical variables. To identify factors associated with ABO, univariable and multivariable logistic regression analyses were performed. In the univariable analysis, explanatory variables with a p value <0.25 were candidates for a multivariable logistic regression model to monitor the influence of confounding variables. With their 95% confidence intervals (95% CI), crude (cOR) and adjusted (aOR) odds ratios were determined to assess the strength and existence of an association. The level of significance α = 0.05 was considered.

Ethics approval and consent to participate
The study complies with the Declaration of Helsinki and was approved and consented to by dedicated ethics oversight bodies such as the Ministry of Health of STP and by the main board of HAM, since at the time the study protocol was submitted, there was no ethics committee in STP. All methods in our study were performed in accordance with the relevant guidelines and regulations in practice. Written informed consent was obtained from all participants after the purpose of the research was explained orally by the researcher. Approval by the participantsṕ arents or legal guardians was asked in the case of adolescents under 16 years of age or for illiterate women. The participants or their legal representatives also consented to have the results of this research work published. Participation in the survey was voluntary, as participants could decline to participate at any time during the study.
The maternal characteristics as well as antepartum, intrapartum, and postpartum factors for all participants and for cases versus controls are described in Table 2.

Factors associated with adverse birth outcomes
In the univariable logistic regression, number of ANC contacts, twin pregnancy, delivery assisted by midwives or obstetricians, preeclampsia, PROM, obstructed labor, cesarean section, meconium-stained amniotic fluid, and infectious risk were eligible for multivariable analysis (Table 3).

Discussion
Assessing neonatal ABOs and identifying contributing factors can help avoid neonatal mortality and morbidity thoroughly and thoughtfully. As a result, the goal of this study was to identify the factors related to ABOs in neonates admitted at birth to HAM in the capital city of    STP. Multiple births (twins), meconium-stained amniotic fluid and PROM were all identified as significant associated factors for ABOs in the current study. This study showed a rate of 17.7% PTB and 16% LBW, which are higher than the published estimates for STP of 12% PTB and 6.6% LBW [2,40,46]. In resource-constrained countries, one explanation for the difference found is the misclassification of late preterm babies (gestational age <37 weeks) in term babies [40,47]. Since gestational age in LMICs is mainly estimated from the date of onset of the last normal menstrual period, most times without ultrasound confirmation, mistakes are frequent [47].
Regarding LBW rates in the country, the source used is the UNICEF Multiple Indicator Cluster Surveys (MICS) [20,21]. MICS data rely upon the information provided by the mother about last birth in the previous two years and are associated with a higher risk of recall bias; therefore, these rate discrepancies between this study and official STP data can be linked to the above reasons. The lack of difference between the rate of PTB and LBW in this study is in line with the studies done in Nepal [48], Ethiopia [49] and Kenya [14] as biologically, a PTB has a higher risk of having LBW as they are less likely to get sufficient time for maturity, growth, and nutrient intake. Another important consideration is that most PTBs in this study were "late preterm" (76/92), often associated with a normal postnatal clinical course with no relevant complications, when compared to very preterm babies (16/92) [50]. The 1.5% congenital malformation and 4% macrosomia rates found in this study are similar to those described by other LMICs [4,8,36].
The neonatal sepsis rate published for LMICs ranges between 1.6% and 3.8% of all live births [50], with disparities among studies regarding the clinical algorithms used and the lack of gold-standard blood cultures for diagnosis [51,52]. This study 4% rate is in line with the overall LMIC rate but neonatal sepsis rates in STP are probably much higher since only babies diagnosed between the 24 th and 36 th hours of life were included; thus, all those with onset of sepsis after being discharged were missed out.
The 8.1% rate of birth asphyxia found in this study is lower when compared to other LMICs: a study in Ethiopia reports a pooled prevalence of 22.52%, 18% in other East African countries and 9.1% in some Central African countries [53,54]. The lower burden of asphyxia in STP may be due to differences in case definition, as in this study, birth asphyxia was only based on a fifth minute APGAR score less than 7, whereas other studies also used other criteria, such as umbilical cord pH < 7-or 20-min Apgar score less than 7 or multiorgan failure in the first 72 h or convulsion in the first 24 h of life [53]. APGAR scoring is also vulnerable to midwife evaluation and therefore susceptible to higher scoring for better health-related outcomes [26,27]. Bold text for p-value �0.05.
Note: Most pregnant women received tetanus toxoid vaccination, iron supplementation and blood pressure measurements, and these factors were not considered. All mothers stated no consanguinity with the baby´s father and did not have smoking habits; therefore, these factors were also not included.
https://doi.org/10.1371/journal.pone.0276348.t002 The fivefold higher risk of multiple pregnancies having ABOs can be associated with the fact that monochorionic pregnancies have a vascular anastomosis within the placenta, affecting the perfusion of each twin and promoting adverse outcomes such as preterm labor, premature rupture of membranes, antepartum hemorrhage and fetal death [55]. Adverse outcomes in twin pregnancies were also reported in other studies [55][56][57]. As an example, some studies report that twin pregnancies have a thirteen-fold increase in rates of stillbirth in monochorionic pregnancies and a fivefold increase in dichorionic twins compared with singleton pregnancies [57]. Hence, the association between multiple pregnancies and ABO in this study reveals the importance of screening for multiple pregnancies as one key component of ANC to reduce the risk of ABO [56]. Other interventions to reduce ABOs related to twin pregnancies that can be recommended in STP are delivery at 37 weeks gestation in uncomplicated dichorionic twin pregnancies and delivery at 36 weeks in monochorionic pregnancies, as proposed by Cheong-See F et al. [57].
In this study, complete ANC with eight or more contacts was a protective factor, which should be expected, since ANC is the most important practice for mothers to obtain more information about nutrition and health, to perform screenings and to learn about danger signs regarding pregnancy and childbirth [16,22]. If a mother lacks ANC, minor obstetric conditions are not detected and managed early; therefore, serious complications and ABOs will likely develop [22,58]. Our finding is similar to the results from Ethiopia and other LMICs, in which ABOs, especially PTB and LBW, were higher among mothers with few ANC contacts [8,18,22,59]. This cutoff is linked to the fact that eight or more ANC contacts can reduce perinatal deaths by up to 8 per 1000 births when compared to only four ANC visits [60].
The rate of pregnancies with meconium-stained amniotic fluid in this study (17.3%) is similar to the international standard and from other studies in LMICs [10,61,62]. The occurrence of meconium-stained amniotic fluid during labor has long been considered a predictor of ABOs and an important sign of fetal distress associated with high rates of neonatal resuscitation, respiratory distress, lower Apgar score, birth asphyxia, neonatal care unit admissions and meconium aspiration syndrome [10,63]. Additionally, approximately 5-10% of neonates with meconium will experience meconium aspiration syndrome, which accounts for approximately 12% of neonatal mortality (as much as a 40% case fatality rate for the neonate and approximately 2% of perinatal mortality) as well as for neonatal sepsis and pulmonary disease, representing an important risk factor not only for ABOs but also for death [10,63]. The odds of experiencing an ABO, in this study, were identified as being approximately twofold higher among meconium-stained fluid compared with a clear amniotic fluid at birth. Hence, early detection by using a latent follow-up chart and partograph and timely intervention is recommended to reduce this significant risk factor [10,63]. PROM was defined as rupture of the membrane lasting more than 18 hours before labor [37,43]. The overall rate in this study was 7.3%, which is in line with the estimated 5%-10% of all pregnancies in LMICs [64,65]. It was also identified as a significant risk factor for ABO, with a threefold higher risk compared to newborns without PROM. This is a well-known risk factor associated with early-onset neonatal sepsis and increased risk for perinatal mortality [64][65][66][67]. Thus, preventive measures should focus on the recognition of these high-risk newborns with early treatment with empirical antibiotics [64][65][66][67]. Such approaches would be a safe and cost-effective strategy, especially in STP where there are no laboratory culture techniques available.
In this study, no association was found between maternal infectious diseases (malaria, HIV, syphilis) and ABOs. This is mainly due to the fact that STP, in contrast to most SSA countries, is about to reach the elimination of HIV, syphilis, hepatitis B and malaria supporting the reduced number of mothers infected enrolled in this study [68,69]. Other viruses known to cause ABOs in SSA countries, such as Zika and dengue were not evaluated in this study since there is no evidence of Zika virus transmission in the country, and the first dengue fever outbreak in STP occurred two years after the completion of the field study [70]. Understanding the role and impact of environmental factors in ABOs is important, and contrary to expectations, no association was found between maternal sanitation behavior (open defecation) or type of access to water (no-improved) and ABOs, as reported in other LMICs studies [15,17,67].

Strengths and limitations
In this study, all clinical pregnancy-related information was abstracted from ANC cards and maternity registers to limit recall bias. Moreover, this study analyzed the main variables that are most frequently associated with ABOs in LMICs, including factors related to i) maternal health conditions, ii) infectious diseases, iii) environmental factors, iv) healthcare access, and v) obstetric interventions.
Regarding the limitations, maternal factors such as gestational weight gain, BMI, and height were not analyzed in this study, as these measurements are not performed during ANC contacts in the country, missing key mediators for ABOs [12]. Participants were asked about the weight gained during the pregnancy or their height, but mothers were not aware of either their height or weight gain. This lack of knowledge of basic one´s features and body conscience might also be due to the lack of scales and stadiometers in most healthcare facilities.
Notwithstanding these limitations, this study represents a significant first step in the description of ABOs and key modifiable associated factors in this resource-constrained setting.

Future research
The scope of this case-control study missed-out some recent areas of research, such as the impact of water contamination, traditional herbs, exposure to toxins on birth outcomes and long-term implications of ABOs on child development, growth, and health outcomes, which should be a matter of future research in this country [15][16][17]. This will promote further understanding of the complex determinants of ABOs and provide new clues on how to improve birth outcomes and maternal and child health.

Conclusions
The high rates of ABOs could be reduced through the provision of high-quality intrapartum care, as twin pregnancy, meconium-stained amniotic fluid and PROM were identified as factors significantly associated with ABOs in this study. Having an ANC with 8 or more contacts was found to be protective, reinforcing the key strategy to reduce ABOs by providing a complete ANC service throughout the continuum of care.
Therefore, the modifiable associated factors found in this case-control study should be considered in cost-effective interventions. Reducing this ABO burden will not only impact neonatal mortality rates but will also promote child well-being, growth, and favorable health outcomes across their life course and provide substantial population-level human capital returns in this small and resource-constrained SSA country.